Stoichiometric Production of Hydrogen Peroxide and Parallel Formation of Ferric Multimers through Decay of the Diferric-Peroxo Complex, the First Detectable Intermediate in Ferritin Mineralization
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- 作者：Guy N. L. Jameson ; Weili Jin ; Carsten Krebs ; Alice S. Perreira ; Pedro Tavares ; Xiaofeng Liu ; Elizabeth C. Theil ; and Boi Hanh Huynh
- 刊名：Biochemistry
- 出版年：2002
- 出版时间：November 12, 2002
- 年：2002
- 卷：41
- 期：45
- 页码：13435 - 13443
- 全文大小：142K
- 年卷期：v.41,no.45(November 12, 2002)
- ISSN：1520-4995
文摘
The catalytic step that initiates formation of the ferric oxy-hydroxide mineral core in thecentral cavity of H-type ferritin involves rapid oxidation of ferrous ion by molecular oxygen (ferroxidasereaction) at a binuclear site (ferroxidase site) found in each of the 24 subunits. Previous investigatorshave shown that the first detectable reaction intermediate of the ferroxidase reaction is a diferric-peroxointermediate, Fperoxo, formed within 25 ms, which then leads to the release of H2O2 and formation offerric mineral precursors. The stoichiometric relationship between Fperoxo, H2O2, and ferric mineralprecursors, crucial to defining the reaction pathway and mechanism, has now been determined. To thisend, a horseradish peroxidase-catalyzed spectrophotometric method was used as an assay for H2O2. Byrapidly mixing apo M ferritin from frog, Fe2+, and O2 and allowing the reaction to proceed for 70 mswhen Fperoxo has reached its maximum accumulation, followed by spraying the reaction mixture into theH2O2 assay solution, we were able to quantitatively determine the amount of H2O2 produced during thedecay of Fperoxo. The correlation between the amount of H2O2 released with the amount of Fperoxo accumulatedat 70 ms determined by Mössbauer spectroscopy showed that Fperoxo decays into H2O2 with a stoichiometryof 1 Fperoxo:H2O2. When the decay of Fperoxo was monitored by rapid freeze-quench Mössbauer spectroscopy,multiple diferric 
-oxo/-hydroxo complexes and small polynuclear ferric clusters were found to form atrate constants identical to the decay rate of Fperoxo. This observed parallel formation of multiple products(H2O2, diferric complexes, and small polynuclear clusters) from the decay of a single precursor (Fperoxo)provides useful mechanistic insights into ferritin mineralization and demonstrates a flexible ferroxidasesite.
-oxo/-hydroxo complexes and small polynuclear ferric clusters were found to form atrate constants identical to the decay rate of Fperoxo. This observed parallel formation of multiple products(H2O2, diferric complexes, and small polynuclear clusters) from the decay of a single precursor (Fperoxo)provides useful mechanistic insights into ferritin mineralization and demonstrates a flexible ferroxidasesite.